Vitamin K is required for carboxylation of specific glutamic acid residues in proteins, converting them to gamma-carboxyglutamic acid (Gla). Gla residues in proteins exhibit specific calcium-binding and calcium-mediated phospholipid binding properties. Anticoagulants such as warfarin inhibit formation of Gla, thereby altering calcium-binding properties of Gla proteins. In addition to liver-derived blood coagulation factors and the well-characterized Gla protein of bone (osteocalcin), other normal tissues and tumor cell types produce Gla proteins of unknown function. In order to understand the mechanism by which warfarin anticoagulants inhibit tumor cell growth in vivo and in vitro, the basic biochemistry of tumor Gla proteins is under investigation. Model cell lines include mouse renal adenocarcinoma (RAG), human osteosarcoma (2T, MG63), and mouse melanoma (B16). The role of Gla proteins in proliferation, motility, and adhesion are under investigation. Warfarin inhibits growth in vitro and alters cellular thiol proteinase enzyme activity as well as generally depressing Gla-protein biosynthesis. Thus, the warfarin-sensitive target may be the cathepsin B-like thiol proteinase, which has been shown to correlate with malignancy in other studies. Recent studies have identified a precursor of this thiol proteinase that is secreted by tumor cells and whose activity is depressed 2-\to 3-fold by warfarin. Purification of this enzyme is in progress. (N)